Valproic acid (Chemical Abstracts Service (CAS) Registry No. 99-66-1) is a branched carboxylic acid having the molecular formula C8H16O2. Valproic acid is also known as 2-propylpentanoic acid, 2-propylvaleric acid, and dipropylacetic acid. Valproic acid is a colorless liquid having a boiling point of 120-121° C. at 14 torr. The compound is very slightly soluble in water. It has a pKa of 4.6, and reacts with bases to form salts generally known as valproates.
Clinical Uses
Since its introduction into the clinical practice in the 1970's, valproic acid (valproate) has been approved by regulatory agencies around the world, including the U.S. Food and Drug Administration (FDA), as a therapy for several clinical indications, including neurological disorders, mania, manic episodes associated with bipolar disorder, convulsions, epilepsy, and affective and attention deficit disorders. In addition, valproate is used for the prophylactic treatment, modulation and management of migraine headache, chronic pain, and neuropathic pain.
Further, potential therapeutic benefits of valproate in still other clinical indications are being evaluated in on-going clinical trials. Valproate therapy is being evaluated in clinical studies assessing activity of the substance as a histone deacetylase inhibitor to promote cell differentiation and regeneration, or to regulate gene expression in subjects afflicted with spinal muscular atrophy. Likewise, valproate may exhibit therapeutic benefit as a combinatorial therapeutic treatment of human cancers and for the treatment of tumor metastasis. Similarly, valproate may be useful in the treatment and management of pain, for treating severe tinnitus, for treatment of disorders of personal attachment and deficient social interaction, or for treating Alzheimer's disease. Pre-clinical studies also show that valproate may promote neural stem cell differentiation and or be useful as a co-medicament to promote the elimination of the Human Immunodeficiency Virus (HIV) or other retroviruses from the body or to prevent progression of a retroviral infection to AIDS.
Although the underlying therapeutic mechanisms are unclear, a growing body of evidence suggests that valproate has neuroprotective and neurotrophic actions. For example, both brain imaging and post-mortem studies demonstrate that bipolar disorder involves a decrease in the volume and number of neurons and glia in discrete brain areas thought to be important for cognition and mood regulation. Remarkably, the reduction in brain volume in bipolar patients was found to be largely suppressed by chronic treatment with valproate, in part as a consequence of its histone deacetylase inhibition. [Kanai H, Saws A, Chen R W, Leeds P, Chuang D M. Valproic acid inhibits histone deacetylase activity and suppresses excitotoxicity-induced GAPDH nuclear accumulation and apoptotic death in neurons. Pharmacogenom J 2004; 4: 336-344.] Likewise, in cellular models, valproate protects rat cerebral cortical neurons and cerebellar granule cells from glutamate-induced excitotoxicity and apoptotic death from stress on the endoplasmic reticulum in C6 glioma cells and PC12 cells. [Bown C D, Wang J F, Chen B, Young L T. Regulation of ER stress proteins by valproate: therapeutic implications. Bipolar Disord 2002; 4: 145-151.] In a rat model of stroke, post-insult valproate treatment reduces ischemia-induced brain damage, caspase-3 activation and neurological deficits. [Ren M, Leng Y, Jeong M, Leeds P R, Chuang D M. Valproic acid reduces brain damage induced by transient focal cerebral ischemia in rats: potential roles of histone deacetylase inhibition and heat shock protein induction. J Neurochem 2004; 89: 1358-1367.] A number of studies report that valproate activates cell survival factors such as Akt, extracellular signal-regulated protein kinase, and cyclic AMP response element binding protein. [De Sarno P, Li X, Jope R S. Regulation of Akt and glycogen synthase kinase-3 beta phosphorylation by sodium valproate and lithium. Neuropharmacology 2002; 43: 1158-1164. Yuan P X, Huang L D, Jiang Y M, Gutkind J S, Manji H K, Chen G. The mood stabilizer valproic acid activates mitogen-activated protein kinases and promotes neurite growth. J Biol Chem 2001; 276: 31674-31683. Einat H, Yuan P, Gould T D, Li J, Du J, Zhang L, et al. The role of the extracellular signal-regulated kinase signaling pathway in mood modulation. J Neurosci 2003; 23: 7311-7316.] Additionally, valproate induces cytoprotective proteins such as Bcl-2, Grp78, brain-derived neurotrophic factor, and heat-shock protein 70. [Chen G, Zeng W Z, Yuan P X, Huang L D, Jiang Y M, Zhao Z H et al. The mood-stabilizing agents lithium and valproate robustly increase the levels of the neuroprotective protein bcl-2 in the CNS. J Neurochem 1999; 72: 879-882.] Moreover, valproate promotes neurite outgrowth. [Yuan P X, Huang L D, Jiang Y M, Gutkind J S, Manji H K, Chen G. The mood stabilizer valproic acid activates mitogen-activated protein kinases and promotes neurite growth. J Biol Chem 2001; 276: 31674-31683.] Recently, valproate was shown to protect dopaminergic neurons in midbrain neuron/glia cultures by stimulating the release of neurotrophic factors from astrocytes. [Chen P-S, Peng G-S, Li G, Yang S, Wu X, Wang C-C, Wilson B, Lu R-B, Gean P-W, Chuang D-M, Hong J-S. Valproate protects dopaminergic neurons in midbrain neuron/glia cultures by stimulating the release of neurotrophic factors from astrocytes. Molec Psych 2006; 11: 1116-1125.] Further, valproate at therapeutic levels was reported to inhibit histone deacetylase (HDAC), an enzyme that catalyzes the remove of acetyl groups from lysine residues of histones, thereby altering gene expression. [Phiel C J, Zhang F, Huang E Y, Guenther M G, Lazar M A, Klein P S. Histone deacetylase is a direct target of valproic acid, a potent anticonvulsant, mood stabilizer, and teratogen. J Biol Chem 2001; 276: 36734-36741. Gottlicher M, Minucci S, Zhu P, Kramer O H, Schimpf A, Giavara S et al. Valproic acid defines a novel class of HDAC inhibitors inducing differentiation of transformed cells. EMBO J. 2001; 20: 6969-6978.]
Sources of the Active Pharmaceutical Ingredient Valproate
Although valproate is a therapeutically active pharmaceutical ingredient, valproic acid is an oil that is difficult to formulate and use in the preparation of dosage forms suitable for human or veterinary use. Pharmaceutical and pharmacological advantages are gained when therapeutic dosage forms are prepared from alkali metal or alkaline earth metal salts of valproic acid. Therefore, alkali metal or alkaline earth metal salts of valproic acid are used in present-day clinical formulations as sources of the active drug ingredient, valproate.
Sodium (Na1+), calcium (Ca2+) and magnesium (Mg2+) valproates have been evaluated for use in pharmaceutical and veterinary compositions. Sodium valproate is a hygroscopic salt that is difficult to formulate into pharmaceutical formulations. In contrast, non-stoichiometric valproate sodium compounds comprising combinations of sodium valproate and valproic acid (divalproex sodium, for example) are not hygroscopic, and are bioavailable and therapeutically active sources of valproate. (The non-stoichiometric compound known as divalproex sodium is disclosed in U.S. Pat. No. 4,988,731, for example, and one of its therapeutic embodiments is described in the FDA Approved Labeling Text for NDA 21-168, Aug. 4, 2000.) At the present time, divalproex sodium is the most commonly formulated source of the drug valproate.
Calcium valproate has also been evaluated for use in pharmaceutical and veterinary formulations. Methods for the preparation of calcium salts of valproic acid are disclosed in U.S. Pat. No. 4,895,873. Although pharmaceutical formulations comprising calcium valproate have been approved by the regulatory bodies of several countries, the use of this valproate salt has been severely restricted following publication of reports of adverse toxicological and reproductive effects in dogs, rats, mice, rabbits, and rats. (For example, adverse effects caused by calcium valproate administration are reported in “Calcium valproate-induced uterine adenocarcinomas in Wistar rats” by Watkins, Gough, et al. in Toxicology, Vol. 41, pages 35-47, 1993.)
Magnesium valproate is also used in clinical formulations. Magnesium valproate, which has the CAS Registry No. 62859-43-7, a molecular formula of C16H30O4Mg, and a molecular weight of 310.71, is also known as magnesium 2-propylvalerate and as 2-propylpentanoic acid magnesium salt. By weight, its composition is 61.8% carbon, 9.7% hydrogen, 7.8% magnesium, and 20.6% oxygen.
Clinical investigators have reported that magnesium valproate possesses pharmacokinetic properties comparable to sodium valproate or valproic acid, is hydrolyzed to valproic acid and magnesium ions upon absorption in the bloodstream, and has important advantages in comparison with either sodium valproate or valproic acid. Among the therapeutic advantages of magnesium valproate are the clinical observations that magnesium valproate exhibits a slower and more regular absorption rate, which prevents the variations in plasma levels of valproate typically observed when sodium salts of valproic acid are administered. Additional therapeutic benefits are afforded by magnesium ions, which possess anticonvulsant and sedative properties. [X. Rabasseda, Drugs of Today, Vol. 31, No. 3, 1995, pp. 185-190.] In contrast to calcium valproate, which exacerbates malignancy, magnesium valproate is a useful therapy when administered to patients with cervical cancer. For example, Chavez-Blanco et al. have reported that magnesium valproate at a dose between 20 and 40 mg/kg inhibits deacetylase activity and hyperacetylates histones in tumor tissues. [A. Chavez-Blanco, B. Segura-Pacheco, et al., Molecular Cancer Jul. 7, 2005, Vol. 4, pp. 22ff.]
Spanish Patent No. ES 430062 discloses one method for the preparation of magnesium valproate in which valproic acid is allowed to react with magnesium oxide in alcoholic medium. The method has the following shortcomings. The reaction is carried out in a suspension. Reaction times are lengthy. The final product is contaminated with unreacted magnesium oxide, which comingles with the desired product when acetone is added to precipitate the magnesium valproate. The resulting product is an amorphous solid that is difficult to purify and dry. The product has poor bioavailability.
In U.S. Pat. No. 5,180,850 to Cavazza, a method is disclosed for the preparation of crystalline magnesium valproate. (The same procedure is disclosed in Italian Patent No. 2,283,789 and in EP 433,848 B1.) According to the method of Cavazza, valproic acid is reacted with a substantially stoichiometric amount of a magnesium alkoxide selected from magnesium ethoxide, magnesium propoxide, and magnesium isopropoxide in methanol or ethanol. The magnesium salt of valproic acid is isolated in a microcrystalline form by solvent evaporation or by acetone precipitation. The method has the following shortcomings. Product isolation by solvent evaporation provides product that is contaminated by incompletely reacted starting materials or adventitious contaminants in starting materials or solvents. When the reaction is carried out in ethanol, the quantity of magnesium ethoxide specified is not completely dissolved in the volume of ethanol taught. Although some conversion to magnesium valproate occurs, the method does not permit control of temperature, reaction time, removal of impurities, etc. When the reaction is carried out in methanol, the addition of acetone fails to precipitate the product, using the volume of acetone taught in the patent.
In U.S. Pat. No. 6,753,349, Weh discloses a method for producing compositions containing at least one molecule of a valproic acid salt and at least one molecule of valproic acid. The valproic acid salt represents an alkali or alkaline earth salt of valproic acid, wherein the alkali salt is a valproate salt of lithium, sodium, potassium, or rubidium, and the alkaline earth salt of valproic acid is a valproate salt of magnesium, calcium, strontium, or barium. Preferably, the valproate salt is a sodium, potassium, magnesium or calcium salt. The compounds of Weh's invention have the general formula:[(CH3CH2CH2)2CH—C(O)O Me]m[(CH3CH2CH2)2CH—C(O)OH]n.xH2Oin which Me is Li1+, Na1+, K1+, Rb1+, Mg2+, Ca2+, Sr2+, or Ba2+, preferably Na1+, K1+, Mg2+, or Ca2+; m is an integer from 1 to 10, preferably from 1 to 6, n is an integer from 1 to 9, preferably from 1 to 3, and the ratio m:n is from 1:1 to 6:1, preferably 1:1 to 5:3 and particularly preferably 1:1, 4:3, or 2:1; and x is zero, 1 or 2, preferably zero or 1. In general, the method of preparing magnesium valproate compositions of Weh's invention comprises combining a selected amount of magnesium carbonate, magnesium bicarbonate, or combinations thereof with a selected amount of valproic acid to form a reaction mixture; and allowing the valproic acid to react directly with the magnesium carbonate, magnesium bicarbonate, or combinations thereof under conditions where the reaction temperature is controlled above the melting point of valproic acid. The methods exhibit the following shortcomings. Neither U.S. Pat. No. 6,753,349 nor related international patents WO 2001/032595 and EP 1,230,205 B1 disclose methods for the preparation of each of the several magnesium valproate compositions that are disclosed in these patents. In the absence of data disclosing the ratios of magnesium carbonate and/or magnesium bicarbonate that must be employed to obtain one of the several magnesium valproate compositions that are disclosed by Weh, a knowledgeable artisan must undertake extensive experiments in order to define a process suitable for pharmaceutical manufacturing. Further, valproic acid is an oil and not a solid with a known melting point, so omission in the disclosure of an optimal reaction temperature also requires extensive experimentation. The final product is contaminated with unreacted magnesium carbonate or bicarbonate, as well as other magnesium valproate salts, all of which comingle with the desired product. No methods for product purification are disclosed. The bioavailability of Weh's magnesium valproate compositions is not reported.
After receiving valproate in its conventional compositions, patients frequently experience deleterious side effects, including gastrointestinal distress and ulceration and occasionally, life-threatening hepatic dysfunction. Hepatotoxicity induced by valproate is characterized by microvesicular periportal steatosis and distorted mitochondria. [H Zimmerman, K Ishak. Valproate-induced hepatic injury: analyses of 23 fatal cases. Hepatology 1982; 2: 591-597.] Studies in animal models and human subjects indicate that following valproate administration, the hepatic levels of free Coenzyme A (CoA) are decreased, and the hepatic levels of medium chain acyl CoA compounds are increased. The co-administration of L-carnitine blocks the development of valproate-induced steatosis. [T P Bohan, P Rogers, C R Roe. Valproate and carnitine. In: Current Concepts in Carnitine Research (A L Carter, Ed.), CRC Press, Boca Raton, Fla., 2000, pp. 19-26.]
L-(−)-Carnitine is a vitamin-like nutrient that is essential for energy production and fat metabolism in the physiological systems of birds, fish, and mammals. The structure of L-carnitine is:

L-carnitine is supplied to the body through both endogenous synthesis (about 25% of the adult daily requirement) and food intake (about 75% of the adult daily requirement). The main dietary source of L-carnitine is meat; beef and lamb provide the most dietary L-carnitine. (Fruits and vegetables provide only traces of L-carnitine.) Within the human body, the major sites of L-carnitine biosynthesis are the liver and kidney, as well as the brain and testes. Biosynthesis requires lysine, methionine, vitamin C, iron, vitamin B6, and niacin. L-carnitine is an essential nutrient for infants, since neonates and young children lack the capacity to synthesize L-carnitine in the quantities that are needed for optimal development.
L-(−)-Carnitine functions as a requisite mediator of acyl transport and accepts acyl groups from a variety of acylCoA derivatives in cells and tissues throughout the body. In humans, the transport activity of L-carnitine is particularly important in working muscle, for example, in the skeletal muscles and the heart. Both types of tissues are dependent on fatty acid metabolism for energy supply, and L-carnitine mediates the translocation of fatty acyl groups across mitochondrial membranes to the sites of oxidation in the mitochondria. In addition, L-carnitine shuttles short chain fatty acids from inside the mitochondria to the cytosol. Other physiological roles of L-carnitine include mitochondrial long-chain fatty acid oxidation, buffering of the mitochondrial acyl CoA/CoA couple, scavenging acyl groups, peroxysomal fatty acid oxidation, branched-chain amino acid oxidation, and membrane stabilization.
Because L-carnitine functions as a requisite mediator of acyl transport in the body, an L-carnitine deficiency is a serious physiological disorder. Individuals who suffer from L-carnitine deficiency are afflicted with muscle weakness (myasthenia), accompanied by an accumulation of lipids in specific types of muscle fibers. Severe L-carnitine deficiency may present as myasthenia gravis. Individuals who suffer from systemic L-carnitine deficiency and also secondary L-carnitine deficiency associated with organic acidemias may experience vomiting, stupor, confusion and in severe or prolonged occasions of systemic L-carnitine deficiency accompanied by stressful stimuli, coma in encephalopathic episodes.
Given the facts that valproate induces liver steatosis and L-carnitine deficiency, that L-carnitine is a requisite mediator of acyl transport in the body, and that neonates and young children lack the ability to synthesize L-carnitine (vide infra), it is not surprising that young age and polytherapy including valproate are the primary risk factors for valproate-induced L-carnitine deficiency and valproate-related hepatic failure. [T P Bohan, P Rogers, C R Roe. Valproate and carnitine. In: Current Concepts in Carnitine Research (A L Carter, Ed.), CRC Press, Boca Raton, Fla., 2000, pp. 19-26.] An incomplete diet, physiological stress situations, such as exercise or pregnancy, and metabolic dysfunction, in particular, lipid disorders or diseases of the liver and kidney, also induce L-carnitine deficiency and create a need for L-carnitine supplementation in adults; this need is increased when valproate is administered.
There are, however, known difficulties in formulating L-carnitine. For example, it is known that L-carnitine is hygroscopic. The hygroscopicity of L-carnitine causes a lack of storability of the solid substance and of simple powder mixtures prepared therefrom, and causes problems such as inadequate flowability during further formulating, processing, and manufacturing of orally administrable dosage forms of pure solid L-carnitine or powdered mixtures containing L-carnitine for use in food, nutritional or dietary supplements for humans or other mammals, animal feed or dietary supplements, or drugs for human or veterinary use. However, oral dosage forms represent the preferred dosage forms, inasmuch as they make it particularly easy for users to take the active ingredient and comply with optimal dosage regimens.
Further, it is known that L-carnitine exhibits a distinctly repugnant malodor and a distinctly objectionable taste after ingestion. The noxious odor and taste render ingestion of oral dosage forms of L-carnitine difficult and interfere with compliance to optimal dosage regimens. Thus, there is a significant unmet need for a form of L-carnitine that is free from noxious odor or taste that can be administered to address L-carnitine deficiency.
Patients receiving conventional valproate compositions often must conform to complex dosing regimens. In addition to valproate, individuals may be taking other medications. Therefore, the addition of a dosage of supplemental L-carnitine often adds a new level of complexity to an already complicated dosing regimen for these patients.
Further, although there are numerous case reports and case series in the literature documenting valproate-induced L-carnitine deficiency, there are very few reports of the carnitine levels in the general patient population. L-Carnitine is not routinely monitored in patients receiving valproate, and unrecognized L-carnitine deficiency may develop unexpectedly in these subjects.
A consideration of all of the facts presented heretofore indicates clearly that there is a significant unmet need for a composition that concomitantly provides therapeutic quantities of both valproate and L-carnitine. The present invention remedies this need.